Activation of ICE-family proteases/caspases initiates apoptosis in mammalian cells. Chemicon's Caspase-3 Colorimetric Activity Assay Kits provide a simple and convenient means for assaying the activity of caspases that recognize the sequence DEVD. The assay is based on spectophotometric detection of the chromophore p-nitroaniline (pNA) after cleavage from the labeled substrate DEVD-pNA. The free pNA can be quantified using a spectrophotometer or a microtiter plate reader at 405 nm. Comparison of the absorbance of pNA from an apoptotic sample with an uninduced control allows determination of the fold increase in caspase-3 activity.

This gene encodes a protein which is a member of the cysteine-aspartic acid protease (caspase) family. Sequential activation of caspases plays a central role in the execution-phase of cell apoptosis. Caspases exist as inactive proenzymes which undergo proteolytic processing at conserved aspartic residues to produce two subunits, large and small, that dimerize to form the active enzyme. This protein cleaves and activates caspases 6, 7 and 9, and the protein itself is processed by caspases 8, 9 and 10. It is the predominant caspase involved in the cleavage of amyloid-beta 4A precursor protein, which is associated with neuronal death in Alzheimer's disease. Alternative splicing of this gene results in two transcript variants that encode the same protein.

FUNCTION: SwissProt: P42574 # Involved in the activation cascade of caspases responsible for apoptosis execution. At the onset of apoptosis it proteolytically cleaves poly(ADP-ribose) polymerase (PARP) at a '216-Asp- -Gly-217' bond. Cleaves and activates sterol regulatory element binding proteins (SREBPs) between the basic helix-loop- helix leucine zipper domain and the membrane attachment domain. Cleaves and activates caspase-6, -7 and -9. Involved in the cleavage of huntingtin.SIZE: 277 amino acids; 31608 Da SUBUNIT: Heterotetramer that consists of two anti-parallel arranged heterodimers, each one formed by a 17 kDa (p17) and a 12 kDa (p12) subunit.SUBCELLULAR LOCATION: Cytoplasm.TISSUE SPECIFICITY: Highly expressed in lung, spleen, heart, liver and kidney. Moderate levels in brain and skeletal muscle, and low in testis. Also found in many cell lines, highest expression in cells of the immune system.PTM: Cleavage by granzyme B, caspase-6, caspase-8 and caspase-10 generates the two active subunits. Additional processing of the propeptides is likely due to the autocatalytic activity of the activated protease. Active heterodimers between the small subunit of caspase-7 protease and the large subunit of caspase-3 also occur and vice versa.SIMILARITY: SwissProt: P42574 ## Belongs to the peptidase C14 family.

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Usage Statement

Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.

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Store kit materials at -20°C up to their expiration date.

Special Precautions:

· After thawing reagents, use immediately or aliquot and freeze at -20oC for longer storage. Avoid repeated freeze/thaw cycles.

· The Caspase-3 substrate and pNA standard are especially light sensitive. Maintain these reagents in amber or covered containers.

Recent evidence suggested that tumor necrosis factor-alpha (TNF-?) and nerve growth factor (NGF) withdrawal activated a common apoptotic pathway. Here, we aimed to investigate the possible role of apoptotic Ras effectors RASSF1 and NORE1 in NGF reduction and TNF-?-related ? cell apoptosis in streptozotocin (STZ)-induced hyperglycemic rats. Rats were divided into four groups: the first group was given saline and citrate buffer, the second group was injected 4-methylcatechol (4-MC), an inducer of NGF synthesis, the third group received STZ, and the fourth group was given both 4-MC and STZ. 4-MC (10 ?g/kg) was administered by daily intraperitoneal injection for 10 days before the animals were rendered hyperglycemic by administration of single dose STZ (75 mg/kg). With 4-MC pretreatment to hyperglycemic rats the following results were noted: (i) Decrease in pancreatic NGF level was blocked, (ii) Increase in pancreatic TNF-? level and the number of TNF-?(+) beta cell in the islets were prevented, (iii) Increase in the number of ? cell synthhesized apoptotic Ras effectors that RASSF1 and NORE1 was blocked, (iv) While pancreatic lipid peroxidation level decreased, antioxidant molecule glutathione and antioxidant enzymes glutathione peroxidase, catalase and superoxide dismutase activities increased, (v) Pancreatic caspase-3 activity and the number of cleaved caspase-3(+) ? cells were decreased. These results strengthen the idea that TNF-? and reduction in NGF can activate a common apoptotic pathway. Moreover, these data display that new apoptotic Ras effector molecules RASSF1 and NORE1 play important role with oxidative stress in NGF reduction and TNF-?-related pancreatic ? cell apoptosis in hyperglycemic rats. Furthermore, these findings suggest that 4-MC can prevent ? cell apoptosis possibly through increasing NGF synthesis in hyperglycemic rats.

Acute right ventricular afterload increase is a known perioperative challenge for the anaesthetic regime especially for patients with a compromised right ventricle. The accused negative inotropic action of volatile anaesthetics, with the exception of xenon, might be crucial for the adaptation of the right ventricle.

Hepatocytes show endoplasmic reticulum (ER) stress when exposed to lipotoxic stimuli such as hyperlipidemia. Recent work has revealed that adenosine monophosphate activated protein kinase (AMPK) can mitigate ER stress. In this study, we investigated the impact of AMPK on lipid-induced ER stress in hepatocytes and its underlying molecular mechanism. Treatment with 5-aminoimidazole-4- carboxamide ribonucleotide (AICAR), an AMPK agonist, or overexpression of a constitutively active AMPK (CA-AMPK) significantly suppressed lipid-mediated ER stress, leading to marked protection against lipotoxic death. Incubation with AICAR and CA-AMPK overexpression induced the expression of an ER-associated chaperone, 150-kDa oxygen-regulated protein (ORP150), at both the mRNA and protein levels in hepatocytes. Forkhead box O1 (FOXO1) was identified as the critical transcription factor regulating ORP150 expression because silencing FOXO1 expression prevented the induction of ORP150 expression by AMPK. In contrast, overexpression of FOXO1-ADA promoted ORP150 expression in hepatocytes. FOXO1 bound directly to the ORP150 promoter, which was enhanced upon in the presence of AICAR. AMPK acts to activate FOXO1 by increasing its deacetylation and transcriptional activity via silent mating type information regulation 2 homolog 1 (SIRT1). Furthermore, AICAR infusion enhanced ORP150 expression, resulting in the marked amelioration of hepatic ER stress and apoptosis in C57BL/6J mice fed a high-fat diet. Our results reveal a novel mechanism by which AMPK regulates ER homeostasis in hepatocytes and suggest that AMPK has a protective role against hypercholesterolemia-related liver damage.

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